Four different methods were used to make visible the nuclei of strains and 2, and They were: (1) Giemsa staining after acid hydrolysis; (2) fluorescence microscopy after acridine orange treatment; (3) acriflavine staining and phase-contrast microscopy; (4) phase-contrast observations with high refractive index mounting medium (polyvinylpyrrolidone). Statistical analyses showed small but significant differences in nuclear counts between the methods. Methods 1 and 2 were somewhat preferable to methods 3 and 4 in that 1 and 2 were easier to score and there was less difference between observers. It was calculated that a single mean nuclear count, by any observer or method, based on 200 scored bacteria should 19 times out of 20 estimate the ‘true’ mean nuclear number of 2 to within about ± 7.5% and of and to within about ± 12%. By increasing the number of bacteria counted and the number of methods used these limits could be appreciably decreased.


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